Amyotrophic lateral sclerosis (ALS) is a devastating and rapidly fatal neurodegenerative disease involving death of upper and lower motor neurons controlling voluntary muscle movement. Current prevalence of ALS in the U.S. is estimated at 20,000, with about 5,000 new cases per year. Though people of all races and ethnicities are equally susceptible to ALS, this disease strikes military veterans more frequently than the general population. Men are also more frequently affected than women. Although ALS is multi-factorial in origin, disease progression and severity uniformly advance as motor neurons die. It is thus expected that neuroprotective agents that block motor neuron death might provide new therapeutic options for patients. However, there are no drugs available that block neuronal cell death, in ALS or any other form of neurodegeneration. Here, we seek to improve the potency, efficacy and safety of the P7C3-class of neuroprotective molecules that we have developed, in hopes of addressing this unmet need. We have previously shown that P7C3-A20, a highly active analog of P7C3, delays motor neuron cell death and loss of motor function in G93A-SOD1 transgenic mice, a preclinical model of ALS. We now propose to evaluate the efficacy of our most highly evolved analogue of P7C3, known as (-)-P7C3-S243, which has shown efficacy in rigorous preclinical models of Parkinson's disease and blast-mediated traumatic brain injury (TBI). Most notably, axonal degeneration is a prominent feature of ALS, and (-)-P7C3-S243 specifically blocks injury- induced axonal degeneration in the absence of neuron cell body death in this model of TBI. We have made substantial progress in medicinal chemistry, and (-)-P7C3-S243 lacks the aniline moiety of the original P7C3 chemical and shows no overt toxicity, including no inhibition of the human hERG channel. Furthermore, prolonged administration of (-)-P7C3-S243 is well tolerated in rodents at doses 10- to 30-fold higher than required for therapeutic efficacy. Importantly, we have also recently identified the molecular target of the P7C3 molecules as nicotinamide phosphoribosyltransferase (NAMPT). NAMPT catalyzes the rate-limiting step in nicotinamide adenine dinucleotide (NAD) salvage, and active analogues of P7C3 enhance its conversion of nicotinamide into nicotinamide mononucleotide (NMN) and NAD in living cells. Strong historical evidence has long predicted that drugs capable of enhancing NAD levels should be uniquely beneficial in treatment of neurodegenerative disease. In addition to mechanistic insight, knowing the molecular target of P7C3 enables us to explore wider swaths of chemistry than previously allowed. Efficacy of new molecules will first be evaluated by in vitro assays of activity, and successful leads will then be evaluated for in vitro and in vivo pharmacokinetic properties. Molecules passing these criteria will be subsequently evaluated in in vivo assays of hippocampal neuroprotection, our original screening platform that identified the P7C3 molecule. Finally, molecules that perform as well or better than our current most promising leads will then be subjected to rigorous testing in two animal models of ALS (G93A-SOD1 mice and ChAT-tTA-9/TDP-43M337V rats), with outcome measures encompassing both motor function and neuronal survival. Protective efficacy in these models will be correlated with CNS levels of the compounds in brain and spinal cord. Our goal is to advance our science from a pre-clinical setting towards first-in-human clinical testing of a neuroprotective drug for ALS.